MECHANISTIC STUDIES RELEVANT TO BROMOURIDINE-ENHANCED NUCLEOPROTEIN PHOTO-CROSS-LINKING - POSSIBLE INVOLVEMENT OF AN EXCITED TYROSINE RESIDUE OF THE PROTEIN

The results of mechanistic studies on formation of uridine (U) and N-a cetyl-m-(5-uridinyl)tyrosine N-ethylamide (2) from irradiation of aque ous, pH 7 solutions of bromouridine (BrU) and N-acetyltyrosine N-ethyl amide (1) are reported. Solutions were irradiated with monochromatic l aser emission at 266, 308 and 325 nm, Quantum yield measurements as a function of excitation wavelength suggest that both products result fr om excitation of the tyrosine derivative followed by electron transfer to BrU, possibly with intermediacy of the hydrated electron. The BrU radical anion ejects bromide to form the uridinyl radical, which then abstracts a hydrogen atom from 1 or adds to the aromatic ring of 1, Fo rmation of adduct 2 is a model for photocrosslinking of nucleic acids bearing the bromouracil chromophore to adjacent tyrosine residues of p roteins in nucleoprotein complexes. The value of 325 nm excitation in photocrosslinking, where the tyrosine chromophore is more competitive for photons, was demonstrated with an RNA bound to the MS2 bacteriopha ge coat protein; more than a 60% increase in the yield of photocrossli nking relative to that obtained with 308 nm excitation was achieved.